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SummaryThe chromosomes of the Gram-positive soil bacteria Streptomyces are linear DNA molecules, usually of about 8 Mb, containing a centrally located origin of replication and covalently bound terminal proteins (which are presumably involved in the completion of replication of the telomeres). The ends of the chromosomes contain inverted repeats of variable lengths. The terminal segments of five Streptomyces chromosomes and plasmids were cloned and sequenced. The sequences showed a high degree of conservation in the first 166-168 bp. Beyond the terminal homology, the sequences diverged and did not generally crosshybridize. The homologous regions contained seven palindromes with a few nucleotide differences. Many of these differences occur in complementary pairs, such that the palindromicity is preserved. Energyoptimized modelling predicted that the 3Ј strand of the terminal palindromes can form extensive hairpin structures that are similar to the 3Ј ends of autonomous parvovirus genomes. Most of the putative hairpins have a GCGCAGC sequence at the loop, with the potential to form a stable single C-residue loop closed by a sheared G:A pairing. The similarity between the terminal structures of the Streptomyces replicons and the autonomous parvoviral genomes suggests that they may share some structural and/or replication features.

The terminal proteins of linear Streptomyces chromosomes and plasmids: a novel class of replication priming proteins tides are similar in length (184-185 amino acids) and sequences, which include a putative helix domain that is homologous to part of the DNA-binding 'thumb' domain of HIV reverse transcriptase, and a putative amphiphilic beta-sheet that may be involved in the observed self-aggregation of the TP and/or the proposed membrane binding. IntroductionSoil bacteria of the genus Streptomyces are unusual among bacteria in having protein-capped linear chromosomes . The terminal sequences of these chromosomes consist of inverted repeats (TIR) of variable lengths ranging from 20 to 550 kb. The nucleotide sequences of these TIR are generally not conserved except for the first ª 200 bp, which are packed with palindromic sequences (Huang et al., 1998a). Streptomyces species also harbour many linear plasmids of a wide range of sizes (tens to hundreds of kilobases). Like the Streptomyces chromosomes, these linear plasmids contain TIR and covalently bound terminal proteins (TPs).Both linear chromosomes and linear plasmids of Streptomyces are replicated from internal origins (Chang and Cohen, 1994;Musialowski et al., 1994). This would leave single-stranded gaps at the 3¢ ends of the replicons (Chang and Cohen, 1994), which are patched by an as yet undefined mechanism presumably involving the TPs (Chen, 1996).The presence of the covalently linked TPs at the telomeres has been indicated by various lines of experimental evidence, including immobilization or retardation of the telomere DNA during electrophoresis in the absence of proteolytic or denaturing treatments (Ito et al., 1978), protection against exonuclease digestion and binding to silica (Coombs and Pearson, 1978;Thomas et al., 1979). The TPs may be readily removed when subjected to mild alkaline conditions, suggesting that the DNA-protein phosphodiester linkage is at a serine residue on the TP. That the TP is linked to the 5¢ end of the DNA has been suggested by protection of the terminal DNA from 5¢ exonucleases but not 3¢ exonucleases, although in at least one case, both 5¢ and 3¢ ends are protected .Linear DNA molecules with 5¢ covalently bound TPs were discovered previously in many eukaryotic and prokaryotic viruses and plasmids, the best studied of which are adenoviruses and bacillus phage f29. These viral genomes have relatively very short (as short as 6 bp in adenoviruses) TIR and are replicated from end to end using the TP as the primer. The TP-primed replication initiation of these viruses contrasts with the proposed TPprimed patching at the Streptomyces telomeres, in that the substrate for the former is double stranded, whereas that for the latter is single stranded. This difference may explain the need for extensive palindromes in the latter, which supposedly fold into elaborate secondary structures containing many hairpin loops and 'bulges' closed by purine:purine sheared pairs resembling those found in the genomes of autonomous parvoviruses (Huang...

Two copies of a DNA sequence similar or identical to one end of the linear plasmid SLP2 were found on the Streptomyces lividans chromosome. Restriction mapping showed that these sequences represented free ends. Electrophoretic retardation and glass-binding studies indicated that the telomeres carry covalently bound proteins. Moreover, the chromosome migrated as an 8 Mb linear DNA in pulsed-field gel electrophoresis. A similar finding with the chromosomes of six other Streptomyces species suggested that a linear chromosome may be characteristic of the genus. The S. lividans chromosome can be circularized by joining the two ends by artificial targeted recombination or by spontaneous deletions spanning both telomeres. Thus the chromosome appears to be able to exist, in viable bacteria, as a linear or a circular molecule.

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